SEASONAL FLUCTUATIONS IN TISSUE NITROGEN,PHOSPHORUS, AND N:P FOR FIVE MACROALGAL SPECIES COMMON TO THE PACIFIC NORTHWEST COAST1

Variations in tissue nitrogen (N) and phosphorus (P) were examined over a complete seasonal cycle for five macroalgae common in Oregon coastal water. Tissue N ranged from 2.0 to 5.5% dry weight (dry wt) in leafy macroalgae (Enteromorpha intestinalis (Linnaeus) Link. Ulva fenestrata Postels et Ruprecht, and Porphyra sp.) and from 0.9 to 2.6% drt wt in branched macroalgae (Codium fragile (Suringar) Hariot and Pelvetiopsis limitata Setchell). Tissue P ranged from 0.32 to 0.86% dry wt in leafy macroalgae and from 0.27 to 0.50% dry wt in branched macroalgae, Ulva fenestrata, C. fragile, and P. limitata appeared to be N limited during part of the year based on tissue N levels. Variations in N:P showed a more distinct seasonal pattern than either tissue N and tissue P. All macroalgae examined appeared to be N limited at least part of the year based on N:P composition, and P limitation occurred for all macroalgae examined except C. fragile. Our results suggest that tissue N:P ratio for macroalgae may be a good index for evaluating in situ nutrient status.     

EFFECTS OF LIGHT ON GROWTH,RuBPCase ACTIVITY AND CHEMICAL COMPOSITION OF ULVA SPECIES (CHLOROPHYTA)1

The effects of light on growth, RuBPCase activity, and chemical composition of Ulva curvata (Kütz.) De Toni and U.Lactuca L. were examined at a range of temperatures and N-supply levels. Groeth of Ulva speices becomes more light-dependent with increasing temperature and N. The effect of light on RuBPcase is N-dependent, with a positive correlation under N-sufficient and a negative correlation under N-limited conditions. Light effects on pigment levels and ratios may be independent of effects on growth rate. These interactions uncouple growth rate from RuBPCase and pigments, and thus from tissue%N. The limits of variability of the growth-%N relationship can be described by a parabola. Under relative light or temperature-limitation, %N is negatively, growth increase with increasing %N. Tight coupling of seaweed wrowth and chemical composition may therefor be relatively rare in natural waters where growth can be simultaneously limited by light, temperature, and N.     

MOLECULAR AND MORPHOLOGICAL ANALYSIS OF ENTEROMORPHA INTESTINALIS AND E. COMPRESSA (CHLOROPHYTA) IN THE BRITISH ISLES

The very common green seaweeds Enteromorpha intestinalis (L.) Nees and E. compressa (L.) Nees are important fouling organisms and have commonly been used as indicators of eutrophication, but their taxonomic status is problematic. The genus presents extreme difficulties because there is wide intraspecific variation in morphology, but morphological differences between species are small and difficult to detect. In this study, molecular data were used in parallel with morphological characters to resolve the taxonomic problems. Phylogenetic analysis of sequences of the internal transcribed spacers ITS1 and ITS2 and the 5.8S gene distinguished two groups of samples, which were identified by morphological characters as E. compressa (branched) and E. intestinalis (normally unbranched). There was a low level of sequence divergence within each group of samples, but divergence between groups was as great as that between either of the two species and the outgroup E. prolifera. Clades representing E. compressa and E. intestinalis were also found in analyses of an independent molecular data set, chloroplast DNA restriction fragment length polymorphisms (RFLPs). Enteromorpha intestinalis and E. compressa represent two distinct, genetically divergent species. Reinterpretation of published studies shows that these species are reproductively isolated. However, E. compressa and E. intestinalis are sometimes very difficult to distinguish from each other and could be regarded as cryptic species. The presence or absence of branching was the most useful character distinguishing these two species, but there was an element of ambiguity because low salinity or salinity shock can induce branching in E. intestinalis. If environmental factors such as salinity are taken into account, branching can be used to identify the great majority of thalli correctly. This study therefore provides a basis for identifying the two most important marine fouling macroalgae and for their use in environmental monitoring and experimentation. Typification of these two Linnaean species showed that current usage of the names accords with the lectotype and protologue of both species. Samples that resembled E. usneoides did not form a clade in any of the trees, and constraining the data to support the monophyly of this group incurred a penalty. Enteromorpha usneoides appears to be an ecotype of E. compressa.     

GROWTH OF ULVA FENESTRATA (CHLOROPHYTA) IN MICROCOSMS RICH IN ZOSTERA MARINA (ANTHOPHYTA) DETRITUS1,2

In laboratory experiments discs cut from thalli of Ulva fenestrata Postels & Ruprecht grew in diameter and biomass (dry weight and ash-free dry weight) more when dead leaves of Zostera marina L. were present than when absent. A maximum increase in dry weight of 500% in 14 days occurred with Zostera present compared with only a 200% increase with no detritus. When NO^?₃ and PO^3-₄ were added the weight of an Ulva disc increased by over 800% when Zostera was present but by only 400% in controls. In general penicillin G (475 units/ml) caused a reduction in algal growth in cultures containing detritus. It is concluded that bacteria on the detritus may compete with algae for nutrients but by releasing unknown substances the bacteria promote the growth of Ulva.     

UNCOUPLING OF VARIOUS MEASURES OF GROWTH IN ULVA ROTUXDATA(CHLOROPHYTA) FOLLOWING A LARGE DECREASE IN IRRADIANCE1

A vegetative clone of the chlorophyte macroalga Ulva rotundata Blid. was maintained in an outdoor continuous flow system and subjected to a large decrease in irradiance. Specific growth rates based on changes in fresh (μ_FW) and dry weight (μ_DW) and surface area (μ_SA) were determined using precut disks over the 24 h following a post-sunset transfer from full sunlight (100% I₀) to 9% I₀ All three measures of growth rate were approximately equivalent for untransferred control plants at either limiting (9%) or saturating (100%)I₀. Transferred disks exhibited μ_FW and μ_SA which were slightly lower than 100%I₀ controls and much higher than 9% I₀ controls; μ_DW was nearly identical for transferred disks and 9% I₀ controls. Cell size was unchanged following transfer, indicating that surface area changes reflected a proportional increase in cell number. Cell division therefore continued at a high rate for one day following transfer of U. rotundata to irradiances which are subsaturating for photosynthesis (indicated by μ_Dw). Starch reserves were largely depleted, and the C/N ratio decreased during this period.     

EFFECTS OF NITROGEN AND PHOSPHORUS ON THE ENDOSYMBIONT LOAD OF RHOPALODIA GIBBA AND EPITHEMIA TURGIDA (BACILLARIOPHYCEAE)1

Diatoms of the family Epithemiaceae possess a unicellular nitrogen-fixing cyanobacterial endosymbiont. We investigated the potential of extracellular nitrogen and phosphorus concentrations to affect the endosymbiont load of Rhopalodia gibba O. Müll, and Epithemia turgida Ehr. in field and culture populations. In a growth chamber experiment, monoclonal cultures of R. gibba were exposed to three levels of nitrate-nitrogen. Nutrient-diffusing substrates were used in a lake environment to create nine microhabitats of varying nitrogen and phosphorus ratios for natural populations of R. gibba and E. turgida. The number of endosymbionts per diatom increased as ambient nitrogen became limiting; mean endosymbiont volume increased as nitrogen increased. The mean endosymbiont surface area: volume ratio decreased with increasing nitrogen. Total endosymbiont volume per diatom (the product of the number of endosymbionts per diatom and their individual biovolumes) did not have a simple response to increasing nitrogen. Phosphorus limitation uncoupled the relationship between endosymbiont load and nitrogen. We suspect that flexibility of the endosymbiont load can reduce the metabolic cost to the diatom if the endosymbionts are dependent on the diatom for a resource.     

EFFECT OF CURRENT VELOCITY ON THE DETACHMENT OF THALLI OF ULVA LACTUCA (CHLOROPHYTA) IN A NEW ZEALAND ESTUARY

Experiments were undertaken in a recirculating flume to determine the relationships among water velocity, thallus area, drag, and the probability of thallus breakage or detachment in the foliose green alga Ulva lactuca L. In all specimens tested to breaking point, thalli detached from their bivalve substrates as a result of stipe breakage rather than in midthallus or by holdfast detachment. There was no relationship between thallus size and drag at which detachment occurred. Rather, the probability of detachment was normally distributed about a mean drag of 0. 70 N (95% confidence limits 0.55–0.85 N). Average breaking stress of stipes was 345 kN.m^-2 (95% cl 250–485 kN.m^-2). Similar results were obtained in field experiments where the horizontal force required to detach thalli was measured directly as 0.93 N (95% cl 0.69–1.15 N). Drag coefficients of plants were not constant with water velocity but increased up to 0.4 m.s^-1, declining exponentially at velocities above this. Empirical relationships were established between coefficient of drag and Reynold's number and, hence, among drag, thallus area and water velocity. These relationships permitted estimation of mean water velocity at which plants of a given area would detach .     

PHOSPHORUS AND NITROGEN NUTRITION IN CHONDRUS CRISPUS (RHODOPHYTA): EFFECTS ON TOTAL PHOSPHORUS AND NITROGEN CONTENT,CARRAGEENAN PRODUCTION,AND PHOTOSYNTHETIC PIGMENTS AND METABOLISM1

The existence of a phenomenon in phosphorus (P) nutrition comparable to the “Neish effect” in nitrogen (N) nutrition (an inverse relation between seawater N enrichment and carrageenan content) was investigated in the temperate red alga Chondrus crispus Stackhouse. Plants were preconditioned for 17 d and then cultured under varying enrichments of P (0, 3, 6, 10, 15 μM P·wk^?1) and a constant N enrichment (53.5 μM N·wk^?1) for 5 wk. Tissue total P, tissue total N, and carrageenan contents were then determined. Identical experiments were performed using C. crispus collected during the fall, winter, spring, and summer seasons. The procedure was repeated using material collected during the following fall season and cultured under constant P (6 μM P·wk^?1) and varying N enrichments (0, 3, 6, 10, 25 μM N·wk^?1). In the fall (P) experiment, carrageenan content was the highest [53.1 ± 0.3% DW (dry weight)], and tissue total P content was the lowest (1.71 ± 0.27 mg P·g DW^?1) in plants that received no P enrichment. Carrageenan content was stable (46.1 ± 1.8% DW) for plants given enrichments of 3 μM P·wk^?1 and greater. Thus, a decrease in carrageenan content, concomitant with an increase in tissue total P content, was observed, but only at tissue total P levels below 2 mg P·g DW^?1. As these levels were always higher than 2 mg P·g DW^?1 in the winter, spring, and summer experiments, carrageenan content remained constant within each season at 46.2 ± 1.3, 43.1 m 0.7, and 44.5 ± 0.6% DW, respectively. Nitrogen enrichment of plants collected in the fall did not affect carrageenan content, which was stable at 49.3 ± 0.9% DW. When these plants were compared with those of the previous fall experiment (6 μM P·wk^?1 and 53.5 μM N·wk^?1), a slight increase in carrageenan content was noted. Thus, at sufficiently high concentration, N also decreased carrageenan content in C. crispus. Phosphorus nutrition had no significant effect on photosynthesis versus irradiance parameters (P_max, α, R_d, I_c, and I_k), the contents of the photosynthetic pigments chlorophyll-a, phycoerythrin (PE), phycocyanin (PC), and allophycocyanin (APC), and the ratios PE:APC and PC:APC. In contrast, N nutrition affected both P_maxand the photosynthetic pigment contents. The data indicate that N limitation reduces the number of phycobilisomes but not their size. The greater reduction in phycobiliprotein than chlorophyll-acontent corroborates the natural bleaching phenomenon regularly observed in C. crispus populations during summer when N levels are generally low in seawater. These results suggest that C. crispus in the temperate waters of the Bay of Fundy may experience N limitation, but P limitation is unlikely.     

GAMETOGENESIS AND GAMETE RELEASE OF ULVA MUTABILIS AND ULVA LACTUCA (CHLOROPHYTA): REGULATORY EFFECTS AND CHEMICAL CHARACTERIZATION OF THE “SWARMING INHIBITOR”1

Gametophytes of Ulva mutabilis Føyn and Ulva lactuca L. were artificially induced to form gametangia by removal of sporulation inhibitors. After this treatment, U. mutabilis gametes were ready for swarming on the third morning after induction, while U. lactuca gametangia needed 1–2 d longer for maturation. Release of gametes of U. lactuca was dependent solely upon exposure to the first light in the morning. Gametangia of U. mutabilis, however, also required sufficient dilution of the swarming inhibitor (SWI). SWI was excreted transiently by both Ulva species early during gametogenesis. While the SWI concentration in U. mutabilis medium remained above the inhibitory concentration until the gametangia were mature, the concentration of U. lactuca‐SWI dropped rapidly below this level. In the presence of sufficient SWI, mature gametes of U. mutabilis remained motionless within the gametangia despite light and open exit pores. However, using SEM, an additional seal was detected within these pores, which probably prevented premature swarming until dilution of SWI and exposure to light. Observations by time lapse microscopy and experiments with the myosin kinase inhibitor BDM suggest that the gametes may be either extruded by the gametangium or leave the exit pore by active gliding motion, driven by a myosin‐like motor protein. The SWIs were purified from both Ulva species, and mass spectral analysis showed their molecular masses (292 Da) were identical.     

EFFECT OF NITROGEN SOURCE ON PLATYMONAS (CHLOROPHYTA) CELL COMPOSITION AND AMINO ACID UPTAKE RATES1

The effect of nitrogen source (nitrate, ammonia and/or amino acids) on cell composition and amino acid uptake rates was examined. Substantial levels of free amino acids accumulated intracellularly with all nitrogen sources used. Ammonia accumulated only when provided in the medium. The presence of ammonia in the medium decreased the intracellular accumulation of free amino acids, especially arginine. Amino acid uptake rates were suppressed by the presence of excess nitrogen, especially ammonia. However, the suppression of uptake did not show any particular relation to the nitrogenous cell composition.     

THE EFFECT OF GROWTH IRRADIANCE ON THE COUPLING OF CARBON AND NITROGEN METABOLISM IN CHAETOMORPHA LINUM (CHLOROPHYTA)

The influence of growth irradiance on the non-steady-state relationship between photosynthesis and tissue carbon (C) and nitrogen (N) pools in Chaetomorpha linum (Muller) Kutzing in response to abrupt changes in external nitrogen (N) availability was determined in laboratory experiments. For a given thallus N content, algae acclimated to low irradiance consistently had a higher rate of light-saturated photosynthesis (P_max normalized to dry weight) than algae acclimated to saturating irradiance; for both treatments, P_max was correlated to thallus N. Both P_max and the photosynthetic efficiency (α_dw) were correlated in C. linum grown at either saturating or limiting irradiance over the range of experimental conditions, indicating that variations in electron transport were coupled to variations in C-fixation capacity despite the large range of tissue N content from 1.1% to 4.8%. Optimizing both α and P_max and thereby acclimating to an intermediate light level may be a general characteristic of thin-structured opportunistic algae that confers a competitive advantage in estuarine environments in which both light and nutrient conditions are highly variable. Nitrogen-saturated algae had the same photosynthesis–irradiance relationship regardless of light level. When deprived of an external N supply, photosynthetic rates did not change in C. linum acclimated to low irradiance despite a two-fold decrease in tissue N content, suggesting that the active pools of chlorophyll and Rubisco remained constant. Both α and P_max decreased immediately and continuously in algae acclimated to high irradiance on removal of the N supply even though tissue N content was relatively high during most of the N-starvation period, indicating a diversion of energy and reductant away from C fixation to support high growth rates. Carbon and nitrogen assimilation were equally balanced in algae in both light treatments throughout the N-saturation and -depletion phases, except when protein synthesis was limited by the depletion of internal N reserves in severely N-starved high-light algae and excess C accumulated as starch stores. This suggests that the ability for short-term adjustment of internal allocation to acquire N andC in almost constant proportions may be especially beneficial to macroalgae living in environments characterized by high variability in light levels and nutrient supply.     

TEMPERATURE DEPENDENCE OF GROWTH AND PHOSPHORUS UPTAKE IN TWO SPECIES OF VOLVOX (VOLVOCALES,CHLOROPHYTA)1

The growth of Volvox globator L. and Volvox aureus Ehr. was measured at five temperatures and nine phosphorus concentrations. Growth rates were hyperbolically related to phosphorus concentrations for all temperatures using a Monod growth model. Optimal growth rates of 1.17 and 1.00 doublings d^?1 were obtained at 20°C for V. globator and V. aureus, respectively. Neither species grew at 5°C. The half-saturation constants for growth, K_s, were lower for V. aureus. Phosphorus uptake by both species was also dependent upon external phosphorus concentrations and temperature. At all temperatures, maximum phosphorus uptake (μmol P colony^?1 min^?1) was similar for both species; however, the half-saturation constants for uptake showed significant differences between the species. Comparisons of the kinetic constants for growth and phosphorus uptake suggest that V. aureus will outcompete V. globator under phosphorus limited, conditions.     

PHOTOSYNTHETIC PROPERTIES OF PROTOPLASTS,AS COMPARED WITH THALLI,OF ULVA FASCIATA (CHLOROPHYTA)1

Protoplasts were prepared from Ulva fasciata Delile, and their photosynthetic performance was measured and compared with that of thalli discs. These protoplasts maintained maximal rates of photosynthesis as high as those of thalli (up to 300 μmol O₂·mg chlorophyll^?1·h^?1) for several hours after preparation and were therefore considered suitable for kinetic studies of inorganic carbon utilization. The photosynthetic K_1/2(inorganic carbon) at pH 6.1 was 3.8 μM and increased to 67, 158, and 1410 μM at the pH values 7.0, 7.9, and 8.9, respectively. Compared with these protoplasts, thalli had a much lower affinity for CO₂ but approximately the same affinity for HCO₃^?. Comparisons between rates of photosynthesis and the spontaneous dehydration of HCO₃^? (at 50 μM inorganic carbon) revealed that photosynthesis of both protoplasts (which lacked apparent activity of extracellular/surface-bound carbonic anhydrase) and thalli (which were only 25% inhibited by the external carbonic anhydrase inhibitor acetazolamide) could not be supported by CO₂ formation in the medium at the higher pH values, indicating HCO₃^? uptake. Since both protoplasts and thalli were sensitive to 4,4′-diisothiocyanostilbene-2,2′-disulfonate, we suggest that HCO₃^? transport was facilitated by the membrane-located anion exchange protein recently reported to function in certain Ulva thalli. These findings suggest that the presence of a cell wall may constitute a diffusion barrier for CO₂, but not for HCO₃^?, utilization under natural seawater conditions.     

INORGANIC CARBON LIMITATION OF PHOTOSYNTHESIS IN ULVA ROTUNDATA (CHLOROPHYTA)1

A computerized oxygen electrode Astern was used to make rapid and accurate measurements of photosynthetic light and dissolved inorganic carbon (DIC) response cures with a macroalga. Ulva rotundata Blid. was grown in an outdoor, continuous flow system in seawater under sunlight or 9% of sunlight at Beaufort, North Carolina. The light compensation points in the shade- and sun-grown plants, measured in seawater, were at photon flux densities (PFDs) of 16 and 27 μmol. Photons·m^?2·s^?1, respectively but the quantum yield of O₂ evolution was not significantly different. Rates of photosynthesis in seawater per unit area of thallus under saturating light and rates of dark respiration were about 1.5-fold higher in sun- than in shade-grown plants. The concentration of DIC in seawater (approximately 2 mM) limited photosynthesis at absorbed PFDs above 60–70 μmol photons·m^?2·s^?1 Addition of 20 mM inorganic carbon had no effect on quantum yield but caused about a 1.5-fold increase in the light-saturated photosynthetic rate in both shade- and sun-grown Ulva. The effect of DIC supplementation was greatest in plants grown in October and least in plants grown in June. The light- and DIC-saturated rate of photosynthesis in seawater was similar to the maximum rate obtained by exposing Ulva to 10% CO₂, in the gas phase. The carbon isotope values (δ¹³C, reflecting the ¹³C/¹²C ratio compared to a standard) of Ulva grown in the same seawater supply were dependent on light and agitation. Samples from Beaufort Inlet were more negative (δ¹³C value, ?20.03‰) than those grown in bright light with agitation (δ¹³C value, ?17.78‰ outdoors; ?17.23‰ indoors), which may indicate DIC supply limited carbon uptake in seawater.     

DIFFERENTIATION OF ULVA MUTABILIS (CHLOROPHYTA) GAMETANGIA AND GAMETE RELEASE ARE CONTROLLED BY EXTRACELLULAR INHIBITORS1

Blade cells of Ulva mutabilis Føyn (Chlorophyta) excrete regulatory factors into their cell walls and into the environment. These factors are essential for the maintenance of the vegetative state. “Sporulation inhibitor-1a” (SI-1a) is a glycoprotein that was isolated from the culture medium of axenic Ulva growing as an undifferentiated callus. This protein was unusually stable to denaturing treatments and showed an extremely high apparent molecular mass (M_r) of 1–4 × 10⁷ daltons estimated by size exclusion chromatography. The glycosylation was not essential for activity. SI-1a suppressed gametogenesis completely at concentrations lower than 10^?14 M. When Ulva developed normally in the presence of their symbiotic bacteria, smaller forms of SI-1 accumulated in the medium (10⁴–10⁶ daltons). Sporulation inhibitors of the same size spectrum and with similar properties were also extracted from crude cell walls of nonaxenic Ulva. A class of different nonprotein sporulation inhibitors (SI-2) of very low M_r and yet unknown structure was isolated from the inner space between the two blade cell layers. Excretion of all SI-1 forms decreased with maturation of the thallus, whereas the overall concentration of SI-2 in the thallus stayed constant throughout the life cycle. The SI-2 affected different Ulva species whereas SI-1 was species-specific. Gametogenesis was induced upon removal of both Sporulation inhibitors from small single-layered fragments of mature blades. After a “determination phase” of 23–46 h, dependent on the time of induction within the cell cycle, the cells became irreversibly committed to differentiation and were no longer susceptible to SI-1 or SI-2. Subsequently, during a 28-h “differentiation phase,” 16 progametes were formed by synchronous genome doublings and cell divisions and differentiated into mature gametes. These became motile and were released from the gametangia when the concentration of a “swarming inhibitor” of low M_r, excreted mainly during the “determination phase,” declined below a threshold concentration. The biochemical properties of these regulatory factors and their effects on gametogenesis and gamete release are described.     

HETEROTROPHIC GROWTH OF ULVA LACTUCA (CHLOROPHYCEAE)1

The effect of external glucose (51 mM) and acetate (13 mM) on growth and photosynthetic capacity of Ulva lactuca L. was tested in laboratory cultures over 41 days in the dark and in dim light (0.9 μmol photons·m^?2·s^?1) at 7–8° C. Glucose and acetate had a significant positive effect on growth rate, chlorophyll content, and quantum yield for discs grown in the dark and in dim light. The carbon gain from heterotrophic uptake was low and only allowed U. lactuca to maintain a specific uptake was low and only allowed U. lactuca to maintain a specific growth rate of 0.005 day^?1 compared to 0.06–0.1 day^?1 at higher light intensities. However, plants with added organic substrate maintained a normal chlorophyll content and were able to photosynthesize whereas control plants lost pigmentation and photosynthetic capability after 41 days in both dim light and darkness, probably because of disorganization of the photosynthetic apparatus. This suggest that the ecological significance of heterotrophic uptake is to allow U. lactuca to survive during prolonged low light conditions with an intact photosynthetic apparatus.     

POLYMORPHIC BEHAVIOR OF ULVA LACTUCA (CHLOROPHYTA) IN AXENIC CULTURE. I. OCCURRENCE OF ENTEROMORPHA-LIKE PLANTS IN HAPLOID CLONES1

An isolate of Ulva lactuca L. was brought into axenic culture in both defined synthetic medium and enriched seawater. Haploid clones were established and followed through several generations in both media. Plants possessing distromatic, partially distromatic or completely tubular blades in cross sections, as well as individuals that were completely distromatic in one area of the blade and tubular in another all developed from the same swarmer population. Variations in basal area morphology also occurred. Progeny from swarmers of any of these morphological types showed a similar mixture of morphological variation, indicating non-mutational variability in phenotypic expression of the blade and basal areas. A mechanism by which such variation may occur without mutation is discussed.     

CHANGES IN INTRACELLULAR NITROGEN POOLS AND FEEDBACK CONTROLS ON NITROGEN UPTAKE IN CHAETOMORPHA LINUM (CHLOROPHYTA)1

Changes in the size of intracellular nitrogen pools and the potential feedback by these pools on maximum N uptake (NH₄⁺ and NO₃^?) rates were determined for Chaetomorpha linum (Müller) Kützing grown sequentially under nutrient-saturating and nutrient-limiting conditions. The size of individual pools in N-sufficient algae could be ranked as residual organic N (RON) comprised mainly of amino acids and amino compounds > protein N > NO₃^? > NH₄⁺ > chlorophyll N. When the external N supply was removed, growth rates remained high and individual N pools were depleted at exponential rates that reflected both dilution of existing pools by the addition of new biomass from growth and movement between the pools. Calculated fluxes between the tissue N pools showed that the protein pool increased throughout the N depletion period and thus did not serve a storage function. RON was the largest storage reserve; nitrate was the second largest, but more temporary, storage pool that was depleted within 10 days. Upon N resupply, the RON pool increased 3 × faster than either the inorganic or protein pools, suggesting that protein synthesis was the rate-limiting step in N assimilation and caused a buildup of intermediate storage compounds. Maximum uptake rates for both NH₄⁺ and NO₃^? varied inversely with macroalgal N status and appeared to be controlled by changes in small intracellular N pools. Uptake of NO₃^? showed an initial lag phase, but the initial uptake of NH₄⁺ was enhanced and was present only when the intracellular NH₄⁺ pool was depleted in the absence of an external N supply. A strong negative correlation between the RON pool size and maximum assimilation uptake rates for both NH₄⁺ and NO₃^? suggested a feedback control on assimilation uptake by the buildup and depletion of organic compounds. Enhanced uptake and the accumulation of N as simple organic compounds or nitrate both provide a temporary mechanism to buffer against the asynchrony of N supply and demand in C. linum.     

PHOSPHORUS LIMITATION AND CARBON METABOLISM IN A UNICELLULAR ALGA: INTERACTION BETWEEN GROWTH RATE AND THE MEASUREMENT OF NET AND GROSS PHOTOSYNTHESIS1

Chlamydomonas reinhardii Dangeard was grown in continuous culture under P limitation at a range of dilution rates. Carbon uptake measurements were performed using double isotope (¹²C/¹⁴C) techniques and the fluxes of carbon in the light and dark were analysed over the range of growth rates. ¹⁴C uptake was shown to be equal to gross photosynthesis only at maximum relative growth rates; at low relative growth rates ¹⁴C uptake approximated net photosynthesis. The altered pattern of C uptake was found to be due to the suppression of dark respiration in the light and the release of ¹⁴C0₂ from respiratory pathways at low relative growth rates. Metabolic channelling of ¹⁴C from photosynthetic pathways to respiratory pathways occurred at low growth rates as the specific activity of the respired CO₂ reached 45% of the input gas mixture. These data are discussed in the light of the controversy concerning the measurement of gross and net photosynthesis in natural populations and in the light of models of ¹⁴C uptake in single celled algae. Existing models are shown to be adequate for high relative growth rates but not for low relative growth rates under P limitation.